Technical Abstract:
In ruminants, development of the female reproductive tract starts before birth, and primordial follicles in the ovaries activate during the third trimester of gestation. Lambs born as twins have lower birth weights and smaller ovarian reserves than lambs born as singletons. We hypothesized that ewe lambs that were twins in utero would have smaller reproductive tracts and differential expression of genes involved in early follicular development. Crossbred ewes (n = 112) were sacrificed at the USMARC abattoir on days 75, 95, or 135 of gestation. Gravid uteri were harvested and fetuses were removed and weighed. The reproductive tract was removed from each ewe fetus (n = 47). The left ovary was fixed for histological evaluation and the right ovary was frozen in liquid nitrogen for gene expression analysis. The uterus was weighed and frozen. Ewe fetuses that were twins weighed less on day 135 of gestation than ewe fetuses that were singletons (P = 0.008); however, there was no difference in uterine weight (P = 0.43) between twins and singletons. At day 135, twins had fewer primordial follicles per histological section than singletons (P = 0.03). Anti-Müllerian Hormone mRNA was not detectable by real-time RT-PCR, and levels of myelocytomatosis viral oncogene homolog (v-MYC) and Indian Hedgehog (IHH) mRNA did not differ between twins and singletons (P > 0.05). There was an increase in v-MYC gene expression from day 75 to day 135 as primordial follicles activated and granulosa cells began to proliferate (P = 0.01). Levels of IHH mRNA increased (P = 0.001) between day 95 and day 135 as follicles transitioned to the early secondary stage. Gestating as a twin decreased fetal weights and decreased the number of primordial follicles but did not alter development of the reproductive tract or expression of developmental genes in the ovaries of ewe fetuses. These results suggest that intrauterine growth restriction due to crowding increases the rate of loss of germ cells during the last trimester.